The long-term goal of this research is to define the mechanism(s) by which progesterone (P) increases prolactin secretion in estrogen (E)-primed nonhuman primates. Prolactin secretion is a useful indicator of serotonin function and deficits in prolactin release are reported for patients with major clinical depression. Therefore, understanding the neural pathway by which ovarian steroids regulate prolactin secretion may reveal important information about the role of ovarian steroids in the neural control of depression. Lactotropes do not contain progestin receptors (PR) and hence the action of P on prolactin is probably mediated by the central nervous system. Our previous work suggests that the action of P on prolactin regulatory neurons is transduced by afferent neurons. We hypothesize that serotonin (5HT) stimulates beta-endorphin and gamma-amino butyric acid neurons which in turn, inhibit the arcuate dopamine neurons resulting in increased prolactin secretion. We showed that E increases the expression of the mRNAs for tryptophan hydroxylase (TPH) and decreases the expression of the mRNA for the serotonin reuptake transporter (SERT), but addition of P had no further effect. However, E decreases 5HT1A autoreceptor mRNA and E+P causes a further decrease in the expression of the 5HT1a autoreceptor. If the changes in mRNA are reflected by changes in protein expression, then together the actions of E and P would increase 5HT synthesis, decrease 5HT reuptake (elevate 5HT in the synapse) and permit 5HT neuronal firing to increase. We also reported a discrepancy between hypothalamic and raphe serotonin levels in guinea pigs compared to TPH mRNA levels in monkeys. That is, the final transmitter product increased with E+P and not E alone, whereas TPH mRNA increased with E and there was no further effect of E+P. This observation raised the possibility that P promoted posttranslational processing of the TPH mRNA. To address this question, TPH protein levels were measured with Western blot and densitometric analysis in guinea pigs treated with E and P. TPH protein levels increased with E and addition of P had no further effect. This result is similar to the results obtained in the examination of TPH mRNA in monkeys. Moreover, there was an increase in serotonin, the final transmitter product, in the hypothalamus with E+P and not with E alone. Thus, there continues to be a discrepancy in the level of serotonin induced by ovarian steroids and the effect of ovarian steroids on the mRNA and protein expression of the rate limiting enzyme in serotonin synthesis. Current studies are focused on the regulation of the 5HT1A postsynaptic receptor and the 5HT2A and 2C receptors in macaque hypothalamus by ovarian steroids.